Analysis of NMDA receptor pore dimensions
MetadataShow full item record
N-methyl-D aspartate receptors (NMDARs) are known to be especially involved in synaptic plasticity and memory formation. NMDARs are highly calcium permeable, have voltage-dependent magnesium block and are inhibited by protons and zinc. There is extensive literature, which sheds light on the permeation properties, and biophysical kinetics of the receptor but very little is known about the contribution of pore dimensions to the ion conduction through the receptor. This is in lieu of the recent evolving idea that dynamic nature of the pore can contribute to receptor permeation properties. NMDARs are tetrameric glutamate gated channels which are composed of two obligatory GluN1 subunits and two either GluN2 (A-D) and/or GluN3 (A-B) subunits. These different subunits provide functional diversity to NMDARs and this is further enhanced by alternative splicing of GluN1 subunit family into eight isoforms (GluN1a/b-4 a/b), which differ in the length of the C-terminal domain. NMDARs are also uniquely known for their selective calcium permeability which is controlled by the extracellularly conserved GluN1 “DRPEER motif” in all functional NMDARs. To increase our mechanistic understanding of functional consequences of regulation of NMDARs, we investigated the effects of GluN1 splice variants and extracellular vestibule i.e. ion selectivity filter (DRPEER motif), on dimensions of the narrowest region of the ion conduction pathway. To do this, we first used physiologically relevant permeant cations i.e. Na + and Ca 2+ to characterize the functional differences in naturally occurring splice variants of GluN1 subunit, which differ in the length of the C-terminal domain and in amino acid substitutions of the DRPEER motif, which are critically involved in calcium permeability. Second, we used organic cations of increasing dimensions as a determinant of the size of the narrow region of the pore. We found that molecular variants of the GluN1 C-terminal domain controlled the channel’s calcium permeability (P Ca /P Na ): GluN1 -1a (~3.1) <3a (~5.0) <2a (~6.41) <4a (~8.08) and diameter in the following order i.e. GluN1 -1a (5.5) <3a (~5.6) <2a (~5.78) <4a (~5.8). Based on these results we propose that regulated splicing of GluN1 subunit controls the unitary current amplitude of NMDARs along with slightly modulating pore sizes. Furthermore, previously it was established in the Popescu lab that D658 and R659 are responsible for altered calcium conductance. We found that this is allosterically modulated by changes in pore size as indicated by differential permeability of organic cations. These data together, could add to the complexity of ion-permeation through NMDA channel pore via modulating dimensions of the pore and affecting the functionality of the receptor. These novel findings also have important implications for understanding the structural and mechanistic regulation of ionic flux through the permeation pathway of NMDARs.